Formation of synaptic connections with appropriate postsynaptic targets is essential for proper nervous system function. We propose to examine the role of environmental cues in guiding the growth cones of developing zebrafish, where we can identify individual neurons and watch their development in living embryos. We propose to examine whether target muscles produce permissive signals that promote elongation of the growth cones of all motoneurons or instructive signals that guide the growth cones of specific identified motoneurons as they pioneer cell-specific pathways to cell-specific targets. We will change the relationship between identified motoneurons and their target muscles by transplanting muscle precursors and motoneurons, and observe the subsequent pathway choices made by the growth cones of labeled, identified motoneurons in living embryos. We propose to compare the pathfinding abilities of two populations of motoneurons that develop at different times, to learn whether their growth cones normally extend to target muscles along pathways previously established by the pioneers. We will ablate the pioneers using laser- irradiation to learn whether their presence is required for proper pathfinding by the later-growing motoneurons.